ORIGINAL RESEARCH—BASIC SCIENCE: Heme Oxygenase vs. Nitric Oxide Synthase in Signaling Mediating Sildenafil Citrate Action

Abstract

ABSTRACT Introduction Heme oxygenase (HO) enzyme catalyzes the rate limiting step in oxidative degradation of heme to biliverdin and carbon monoxide (CO). CO has been shown to share many properties with nitric oxide (NO), including activation of guanyl cyclase, signal transduction, and gene regulation. Aim To assess the signaling pathways mediating cavernous tissues response to sildenafil citrate intake experimentally. Main Outcome Measures In dissected cavernous tissues; detection of HO-1, HO-2 and nueronal nitric oxide synthase (nNOS) gene expressions by reverse transcriptase polymerase chain reaction (RT-PCR), HO enzyme activity assay, HO-1, HO-2 protein detection by Western blot, cyclic guanosine monophosphate (cGMP) tissue levels by enzyme linked immunosorbent assay (ELISA) and histopathology. Methods Two hundred forty Sprague-Dawley rats divided into five equal groups were investigated: group (Gr) 1, controls received regular diet; Gr 2, received sildenafil citrate 4 mg/kg orally; Gr 3, received the same dose of sildenafil added to HO inducer, diferuloylmethane; Gr 4, received sildenafil added to HO inhibitor, zinc protoporphyrin, and Gr 5, received sildenafil kg orally by gastric tube. Gr 3 received the same dose of sildenafil added to HO inducer, added to nitric oxide synthase inhibitor, L-Nitroarginine methylester. Twelve rats from each group were sacrificed by cervical dislocation successively after 1/2, 1, 2, and 3 hours from the intake. Results HO-2 gene expression was demonstrated in all groups. HO-1 was not expressed in controls, expressed in Gr 2, accentuated in Gr 3, and attenuated in Gr 4 and 5. These results were confirmed by Western blot. The nNOS was expressed in controls, increased in Gr 2 and 3, and decreased in Gr 4 and 5. HO enzyme activity and cGMP levels were significantly elevated in Gr 2, accentuated in Gr 3, and significantly decreased in Gr 4 and 5 compared to controls. Vasodilatations were observed in cavernous tissues of histopathologic sections of Gr 2 and increased in those of Gr 3. Conclusion Sildenafil citrate actions may be mediated by up-regulation of HO-1 gene expression.